IC4R006-Epigenomic-2016-22110044 - Revision history

Xysj1990: /* Research Findings */

2016-06-22T08:22:45Z

‎Research Findings

Xysj1990: /* Plant Materials & Treatment */

2016-06-22T08:19:27Z

‎Plant Materials & Treatment

Xysj1990: /* The Background of This Project */

2016-06-22T04:42:49Z

‎The Background of This Project

Xysj1990: /* Plant Materials & Treatment */

2016-06-22T04:42:30Z

‎Plant Materials & Treatment

Xysj1990: /* Plant Materials & Treatment */

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‎Plant Materials & Treatment

Xysj1990: /* The Background of This Project */

2016-06-22T04:42:08Z

‎The Background of This Project

Xysj1990: /* The Background of This Project */

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‎The Background of This Project

Xysj1990: /* The Background of This Project */

2016-06-22T04:38:43Z

‎The Background of This Project

Xysj1990 at 04:37, 22 June 2016

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Xysj1990: /* Project Title */

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‎Project Title

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 ==Research Findings==
 * '''DH sites are significantly associated with conserved noncoding sequences (CNSs) and protein-binding cis elements.''' CNSs have been identified in both mammalian and plant species and are significantly associated with regulatory sequences (Freeling and Subramaniam 2009; Haeussler and Joly 2011). We were interested in the association between CNS and the DH site since both types of DNA sequences are likely related to regulatory elements. The liguleless1 gene was extensively studied in grass species, and a total of seven CNSs were identified within this gene (Kaplinsky et al. 2002). A DH site, showing in both seedling and callus tissues, was identified in the 59 UTR of this gene. This DH site is partially overlapped with one of the CNSs (P = 0.034, binomial test) (Fig. 2A).
 * The researchers then examined the association of rice DH sites with the 46,355 CNSs conserved between the rice and sorghum genomes (Schnable et al. 2011). These CNSs span a total of 1.6 Mb of rice genome sequences. We found that 25.7% (11,911) and 41.6% (19,281) of the CNSs were associated with DH sites in the seedling and callus, respectively (P < 0.001, binomial test) (for an example region, see Fig. 2B).
 * Many regulatory proteins bind to specific DNA motives. The PCF1 and PCF2 proteins in rice, which are essential for regulating meristematic tissue-specific expression of rice genes, bind two cis elements (element IIa, AGGTGGGCCCGT, and element IIb, TGTGGGACCATG) (Kosugi and Ohashi 1997). Introducing two mutated bases in element IIa (AGGTGGGCGAGT) resulted in loss of binding affinity to both proteins (Kosugi and Ohashi 1997). The researchers identified a total of 110 regions that have 100% match to the two cis elements in the rice genome. Most of these regions were located outside of genes (82 of 110) or in introns (16 of 110). DH sites were associated with 43 (39%) of these regions (33 of 57 element IIa and 10 of 53 element IIb). Almost all of these 43 DH sites were located in outside of genes (37) or in introns (3). In contrast, we identified 22 regions with 100% match to the AGGTGGGCGAGT sequence. DH sites were associated with only two (9%) of these regions (P < 0.001, binomial test).
 ==Labs working on this Project==

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 ==Plant Materials & Treatment==
 * Mixed leaf and stem tissues of 2-wk-old rice cultivar ''Nipponbare'' seedlings grown in a greenhouse were collected and ground into a fine powder in liquid nitrogen. The resulting powder was suspended in nuclear isolation buffer (NIB; 20 mM Tris-HCl, 50 mM EDTA, 5 mM Spermidine, 0.15 mM Spermine, 0.1% mercaptoethanol, 40% Glycerol at pH 7.5) and followed the standard protocol for nuclei isolation. Rice callus tissue was induced from sterilized ''Nipponbare'' seeds in rice calli induction medium (NB basal medium plus vitamin, glutamine, proline, casein hydrolysate, sucrose, and phytogel as well as 3 mg/L 2,4-D at pH 5.8) under 28°C–29°C with dark conditions. Three-week-old calli were col- lected for nuclei isolation using the same method as for leaf tissue. The prepared nuclei pellet was suspended in RSB buffer (10 mM Tris at pH 7.4, 10 mM NaCl, 3 mM MgCl 2 ) for DNase I (Roche) digestion with increasing concentrations (0–4 units) for 10 min at 37°C.
 ==Labs working on this Project==

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 * The identification and functional characterization of the regula- tory DNA elements is essential for understanding the regulation of gene expression in eukaryotic genomes. Although the genomes of an increasing number of eukaryotic species have been sequenced, genome-wide identification of regulatory DNA elements, such as that being done in the ENCODE project (The ENCODE Project Consortium 2007) and the Epigenomics Roadmap (Bernstein et al. 2010) in humans and in the modENCODE projects in Caenorhabditis elegans and Drosophila melanogaster (Gerstein et al. 2010; Roy et al. 2010), has been initiated only in few species. Active regulatory DNA elements, such as promoter and enhancers, in- teract with regulatory proteins. As a result, these regions are either free of nucleosomes or are under dynamic nucleosome modifications or displacements (Henikoff et al. 2009; Jin et al. 2009). Thus, active DNA elements are associated with ''open chromatin'' in higher eukaryotic genomes. One distinct characteristic of the genomic regions of open chromatin is a pronounced sensitivity to cleavage of endonuclease DNase I (Wu 1980; Keene et al. 1981; McGhee et al. 1981). Almost all active regulatory elements, in- cluding promoters, enhancers, suppressors, insulators, and locus control regions, have been shown to be marked by DNase I hypersensitive (DH) sites. (Gross and Garrard 1988).
 * Rice (Oryza sativa) is the most important food crop in the world and has also been established as a model species for plant genome research. Rice provides one of the most accurately sequenced genomes from any multicellular eukaryotes (Goff et al. 2002; Matsumoto et al. 2005). Extensive genome-wide DNA methylation and histone modification data sets have recently been generated in rice (Feng et al. 2010; He et al. 2010; Yan et al. 2010; Zemach et al. 2010).
-* ''In this project , the researchers describe high-resolution maps of DH sites in rice from both seedling and callus tissues. We report a number of novel features associated with rice DH sites, including their epigenetic modifica- tions, dynamic response to tissue culture, and association with genes that differentially expressed genes in seedling and callus tissues.''
+* '''In this project , the researchers describe high-resolution maps of DH sites in rice from both seedling and callus tissues. We report a number of novel features associated with rice DH sites, including their epigenetic modifica- tions, dynamic response to tissue culture, and association with genes that differentially expressed genes in seedling and callus tissues.'''
 ==Plant Materials & Treatment==

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 ==Plant Materials & Treatment==
-* Mixed leaf and stem tissues of 2-wk-old rice cultivar ‘‘Nipponbare’’ seedlings grown in a greenhouse were collected and ground into a fine powder in liquid nitrogen. The resulting powder was suspended in nuclear isolation buffer (NIB; 20 mM Tris-HCl, 50 mM EDTA, 5 mM Spermidine, 0.15 mM Spermine, 0.1% mercaptoethanol, 40% Glycerol at pH 7.5) and followed the standard protocol for nuclei isolation. Rice callus tissue was induced from sterilized ''Nipponbare'' seeds in rice calli induction medium (NB basal medium plus vitamin, glutamine, proline, casein hydrolysate, sucrose, and phytogel as well as 3 mg/L 2,4-D at pH 5.8) under 28°C–29°C with dark conditions. Three-week-old calli were col- lected for nuclei isolation using the same method as for leaf tissue. The prepared nuclei pellet was suspended in RSB buffer (10 mM Tris at pH 7.4, 10 mM NaCl, 3 mM MgCl 2 ) for DNase I (Roche) digestion with increasing concentrations (0–4 units) for 10 min at 37°C.
+* Mixed leaf and stem tissues of 2-wk-old rice cultivar ''Nipponbare'' seedlings grown in a greenhouse were collected and ground into a fine powder in liquid nitrogen. The resulting powder was suspended in nuclear isolation buffer (NIB; 20 mM Tris-HCl, 50 mM EDTA, 5 mM Spermidine, 0.15 mM Spermine, 0.1% mercaptoethanol, 40% Glycerol at pH 7.5) and followed the standard protocol for nuclei isolation. Rice callus tissue was induced from sterilized ''Nipponbare'' seeds in rice calli induction medium (NB basal medium plus vitamin, glutamine, proline, casein hydrolysate, sucrose, and phytogel as well as 3 mg/L 2,4-D at pH 5.8) under 28°C–29°C with dark conditions. Three-week-old calli were col- lected for nuclei isolation using the same method as for leaf tissue. The prepared nuclei pellet was suspended in RSB buffer (10 mM Tris at pH 7.4, 10 mM NaCl, 3 mM MgCl 2 ) for DNase I (Roche) digestion with increasing concentrations (0–4 units) for 10 min at 37°C.
 ==Labs working on this Project==

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 ==Plant Materials & Treatment==
-* Mixed leaf and stem tissues of 2-wk-old rice cultivar ‘‘Nipponbare’’ seedlings grown in a greenhouse were collected and ground into a fine powder in liquid nitrogen. The resulting powder was suspended in nuclear isolation buffer (NIB; 20 mM Tris-HCl, 50 mM EDTA, 5 mM Spermidine, 0.15 mM Spermine, 0.1% mercaptoethanol, 40% Glycerol at pH 7.5) and followed the standard protocol for nuclei isolation. Rice callus tissue was induced from sterilized
+* Mixed leaf and stem tissues of 2-wk-old rice cultivar ‘‘Nipponbare’’ seedlings grown in a greenhouse were collected and ground into a fine powder in liquid nitrogen. The resulting powder was suspended in nuclear isolation buffer (NIB; 20 mM Tris-HCl, 50 mM EDTA, 5 mM Spermidine, 0.15 mM Spermine, 0.1% mercaptoethanol, 40% Glycerol at pH 7.5) and followed the standard protocol for nuclei isolation. Rice callus tissue was induced from sterilized ''Nipponbare'' seeds in rice calli induction medium (NB basal medium plus vitamin, glutamine, proline, casein hydrolysate, sucrose, and phytogel as well as 3 mg/L 2,4-D at pH 5.8) under 28°C–29°C with dark conditions. Three-week-old calli were col- lected for nuclei isolation using the same method as for leaf tissue. The prepared nuclei pellet was suspended in RSB buffer (10 mM Tris at pH 7.4, 10 mM NaCl, 3 mM MgCl 2 ) for DNase I (Roche) digestion with increasing concentrations (0–4 units) for 10 min at 37°C.
 ==Labs working on this Project==

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 ==The Background of This Project==
-*Cytosine DNA methylation is a conserved epigeneti silencing mechanism in higher eukaryotes. Cytosine methylation plays an important role in many biological processes, including defense against transposon proliferation (Tsukahara et al., 2009), control of genomic imprinting (Morison et al., 2005) and regulation of gene expression (Bird, 2002). In mammals, cytosine methylation is controlled by the de novo methyltransferases DNMT3a/b, and is maintained by the methyltransferase DNMT1 (Goll and Bestor, 2005). Methylated cytosines occur almost exclu- sively at CG dinucleotides in mammalian genomes. However, Lister et al. (2009) recently showed that approximately 15% of methylated cytosines are associated with neighbor.
+* The identification and functional characterization of the regula- tory DNA elements is essential for understanding the regulation of gene expression in eukaryotic genomes. Although the genomes of an increasing number of eukaryotic species have been sequenced, genome-wide identification of regulatory DNA elements, such as that being done in the ENCODE project (The ENCODE Project Consortium 2007) and the Epigenomics Roadmap (Bernstein et al. 2010) in humans and in the modENCODE projects in Caenorhabditis elegans and Drosophila melanogaster (Gerstein et al. 2010; Roy et al. 2010), has been initiated only in few species. Active regulatory DNA elements, such as promoter and enhancers, in- teract with regulatory proteins. As a result, these regions are either free of nucleosomes or are under dynamic nucleosome modifications or displacements (Henikoff et al. 2009; Jin et al. 2009). Thus, active DNA elements are associated with ''open chromatin'' in higher eukaryotic genomes. One distinct characteristic of the genomic regions of open chromatin is a pronounced sensitivity to cleavage of endonuclease DNase I (Wu 1980; Keene et al. 1981; McGhee et al. 1981). Almost all active regulatory elements, in- cluding promoters, enhancers, suppressors, insulators, and locus control regions, have been shown to be marked by DNase I hypersensitive (DH) sites. (Gross and Garrard 1988).
 ==Labs working on this Project==

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 ==Labs working on this Project==
-* Department of Horticulture, University of Wisconsin-Madison, Madison, WI 53706, USA, and
+* Department of Horticulture, University of Wisconsin–Madison, Madison, Wisconsin 53706, USA;
-* U.S. Department of Energy Joint Genome Institute, Walnut Creek, CA 94598, USA
+* Department of Plant and Microbial Biology, University of California–Berkeley, Berkeley, California 94720, USA;
 ==Corresponding Author==
 * '''Jiming Jiang''' (jjiang1@wisc.edu)

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 ==Project Title==
-* '''Genome-wide mapping of cytosine methylation revealed dynamic DNA methylation patterns associated with genes and centromeres in rice'''
+* '''High-resolution mapping of open chromatin in the rice genome'''
 ==The Background of This Project==
 *Cytosine DNA methylation is a conserved epigeneti silencing mechanism in higher eukaryotes. Cytosine methylation plays an important role in many biological processes, including defense against transposon proliferation (Tsukahara et al., 2009), control of genomic imprinting (Morison et al., 2005) and regulation of gene expression (Bird, 2002). In mammals, cytosine methylation is controlled by the de novo methyltransferases DNMT3a/b, and is maintained by the methyltransferase DNMT1 (Goll and Bestor, 2005). Methylated cytosines occur almost exclu- sively at CG dinucleotides in mammalian genomes. However, Lister et al. (2009) recently showed that approximately 15% of methylated cytosines are associated with neighbor.